SUMMARY: Hydrogen-Deuterium Exchange Mass Spectrometry (HDX-MS) monitors deuterium uptake at the peptide level, in a time-dependent manner. It produces complex, multi-dimensional data that must be interpreted at minimum both the temporal and sequence levels. Specialized tools are therefore essential to preprocess, integrate, and analyze HDX-MS data and translate it into meaningful biological insights. HaDeX2 provides statistical inferences and their visualizations across five dimensions of HDX-MS data: protein sequence, time, biological states, peptide charge and experimental replicates. AVAILABILITY AND IMPLEMENTATION: HaDeX2 is freely available as an R package (https://github.com/hadexversum/HaDeX2; https://doi.org/10.5281/zenodo.18543703) and web server (https://hadex2.mslab-ibb.pl/). To run the GUI locally, users should install a dedicated companion package (https://github.com/hadexversum/HaDeXGUI).

MOTIVATION: Hydrogen-deuterium mass spectrometry (HDX-MS) is a rapidly developing technique for monitoring dynamics and interactions of proteins. The development of new devices has to be followed with new software suites addressing emerging standards in data analysis. RESULTS: We propose HaDeX, a novel tool for processing, analysis and visualization of HDX-MS experiments. HaDeX supports a reproducible analytical process, including data exploration, quality control and generation of publication-quality figures. AVAILABILITY AND IMPLEMENTATION: HaDeX is available primarily as a web-server (http://mslab-ibb.pl/shiny/HaDeX/), but its all functionalities are also accessible as the R package (https://CRAN.R-project.org/package=HaDeX) and standalone software (https://sourceforge.net/projects/HaDeX/). SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.